Transcriptome Level Research Articles

HER2-Low Luminal Breast Carcinoma Is Not a Homogenous Clinicopathological and Molecular Entity.

With the development of some new antibody-drug conjugates, the HER2 classification of breast carcinomas now includes the HER2-low (H2L) category: IHC 1+, 2+ non-amplified by ISH, and double-equivocal carcinomas, mostly luminal, expressing hormone receptors (HR+). We analyzed mutational status and transcriptomic activities of three HER2 effector pathways: PI3K-AKT, MAPK, and JAK-STAT, in association with clinicopathologic features, in 62 H2L carcinomas compared to 43 HER2-positive and 20 HER2-negative carcinomas, all HR+. H2L carcinomas had significantly lower histoprognostic grades and mitotic and Ki67 proliferation indexes than HER2-positive carcinomas. Their PIK3CA mutation rates were close to those of HER2-negative and significantly higher than in HER2-positive carcinomas, contrary to TP53 mutations. At the transcriptomic level, we identified three distinct groups which did not reflect the new HER2 classification. H2L and HER2-negative carcinomas shared most of clinicopathological and molecular characteristics, except HER2 membrane expression (mRNA levels). The presence of a mutation in a signaling pathway had a strong pathway activation effect. PIK3CA mutations were more prevalent in H2L carcinomas, leading to a strong activation of the PI3K-AKT signaling pathway even in the absence of HER2 overexpression/amplification. PIK3CA mutations may explain the failure of conventional anti-HER2 treatments, suggesting that new antibody-drug conjugates may be more effective.

Identification and clinical validation of diverse cell-death patterns-associated prognostic features among low-grade gliomas

Low-grade glioma (LGG) is heterogeneous at biological and transcriptomic levels, and it is still controversial for the definition and typing of LGG. Therefore, there is an urgent need for specific and practical molecular signatures for accurate diagnosis, individualized therapy, and prognostic evaluation of LGG. Cell death is essential for maintaining homeostasis, developing and preventing hyperproliferative malignancies. Based on diverse programmed cell death (PCD) related genes and prognostic characteristics of LGG, this study constructed a model to explore the mechanism and treatment strategies for LGG cell metastasis and invasion. We screened 1161 genes associated with PCD and divided 512 LGG samples into C1 and C2 subtypes by consistent cluster analysis. We analyzed the two subtypes' differentially expressed genes (DEGs) and performed functional enrichment analysis. Using R packages such as ESTIMATE, CIBERSOTR, and MCPcounter, we assessed immune cell scores for both subtypes. Compared with C1, the C2 subtype has a poor prognosis and a higher immune score, and patients in the C2 subtype are more strongly associated with tumor progression. LASSO and COX regression analysis screened four characteristic genes (CLU, FHL3, GIMAP2, and HVCN1). Using data sets from different platforms to validate the four-gene feature, we found that the expression and prognostic correlation of the four-gene feature had a high degree of stability, showing stable predictive effects. Besides, we found downregulation of CLU, FHL3, and GIMAP2 significantly impairs the growth, migration, and invasive potential of LGG cells. Take together, the four-gene feature constructed based on PCD-related genes provides valuable information for further study of the pathogenesis and clinical treatment of LGG.

Comparison of the Effect of Pruning on Plant Growth and Transcriptome Profiles in Different Tea Varieties

Although pruning contributes to the growth and development of new shoots, it is important to note that the growth potential and yield of tea varieties may differ after pruning due to genetic and environmental factors. In this experiment, 20 different varieties of tea plants were used to observe their potential for growth, shoot development, and other phenotypic indexes after pruning. The study aimed to determine the suitability of each variety for heavy pruning. It was concluded that there were obvious differences in tree strength and new growth after pruning of the different varieties, with ‘Zhongcha 302’ exhibiting the strongest growth and ‘Emei Wenchun’ showing the weakest growth. In order to understand the molecular mechanisms involved, a transcriptomic analysis was carried out on the two tea varieties. The results of the data indicate that the expression of CYP450 family was high in ‘Zhongcha 302’. In ‘Emei Wenchun’, the expression of NCED was higher than that in ‘Zhongcha 302’. The KEGG results indicate that pruning stimulates the expression of genes involved in phytohormone signalling and plant–pathogen interaction pathways in tea plants. The study offered scientific guidance for tea plant pruning suitability and preliminarily revealed the regulatory mechanism of new shoot growth in different tea plant varieties at the transcriptome level.

Integrative analysis of transcriptome, DNA methylome, and chromatin accessibility reveals candidate therapeutic targets in hypertrophic cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is the most common inherited heart disease and is characterized by primary left ventricular hypertrophy usually caused by mutations in sarcomere genes. The mechanism underlying cardiac remodeling in HCM remains incompletely understood. An investigation of HCM through integrative analysis at multi-omics levels will be helpful for treating HCM. DNA methylation and chromatin accessibility, as well as gene expression, were assessed by nucleosome occupancy and methylome sequencing (NOMe-seq) and RNA-seq, respectively, using the cardiac tissues of HCM patients. Compared with those of the controls, the transcriptome, DNA methylome, and chromatin accessibility of the HCM myocardium showed multifaceted differences. At the transcriptome level, HCM hearts returned to the fetal gene program through decreased sarcomeric and metabolic gene expression and increased extracellular matrix gene expression. In the DNA methylome, hypermethylated and hypomethylated differentially methylated regions were identified in HCM. At the chromatin accessibility level, HCM hearts showed changes in different genome elements. Several transcription factors, including SP1 and EGR1, exhibited a fetal-like pattern of binding motifs in nucleosome-depleted regions in HCM. In particular, the inhibition of SP1 or EGR1 in an HCM mouse model harboring sarcomere mutations markedly alleviated the HCM phenotype of the mutant mice and reversed fetal gene reprogramming. Overall, this study not only provides a high-precision multi-omics map of HCM heart tissue but also sheds light on the therapeutic strategy by intervening in the fetal gene reprogramming in HCM.

Metabolic pathway-based subtypes associate glycan biosynthesis and treatment response in head and neck cancer

Head and Neck Squamous Cell Carcinoma (HNSCC) is a heterogeneous malignancy that remains a significant challenge in clinical management due to frequent treatment failures and pronounced therapy resistance. While metabolic dysregulation appears to be a critical factor in this scenario, comprehensive analyses of the metabolic HNSCC landscape and its impact on clinical outcomes are lacking. This study utilized transcriptomic data from four independent clinical cohorts to investigate metabolic heterogeneity in HNSCC and define metabolic pathway-based subtypes (MPS). In HPV-negative HNSCCs, MPS1 and MPS2 were identified, while MPS3 was enriched in HPV-positive cases. MPS classification was associated with clinical outcome post adjuvant radio(chemo)therapy, with MPS1 consistently exhibiting the highest risk of therapeutic failure. MPS1 was uniquely characterized by upregulation of glycan (particularly chondroitin/dermatan sulfate) metabolism genes. Immunohistochemistry and pilot mass spectrometry imaging analyses confirmed this at metabolite level. The histological context and single-cell RNA sequencing data identified the malignant cells as key contributors. Globally, MPS1 was distinguished by a unique transcriptomic landscape associated with increased disease aggressiveness, featuring motifs related to epithelial-mesenchymal transition, immune signaling, cancer stemness, tumor microenvironment assembly, and oncogenic signaling. This translated into a distinct histological appearance marked by extensive extracellular matrix remodeling, abundant spindle-shaped cancer-associated fibroblasts, and intimately intertwined populations of malignant and stromal cells. Proof-of-concept data from orthotopic xenotransplants replicated the MPS phenotypes on the histological and transcriptome levels. In summary, this study introduces a metabolic pathway-based classification of HNSCC, pinpointing glycan metabolism-enriched MPS1 as the most challenging subgroup that necessitates alternative therapeutic strategies.

Current state of in situ gene expression studiesin animal tissues

Morphological studies of farm animals are most often conducted using simple preparation and staining techniques. The study of the processes of embryogenesis, postembryonic features of the development of organs and tissues, as well as the effect of various substances remains to be elucidated not only using histochemical and immunohistochemical staining methods, but also using RNA in situ hybridization and transcriptome in situ sequencing. Aspects of many cellular and tissue processes for cattle, pigs and chickens in the context of comparative physiology have not yet been studied. The high productivity of farm animals is associated with the intensive functioning of all organs and systems of the body. The influence of the agricultural management of farm animals and its consequences on the development of an organism in ontogeny deserve a separate area of research from the point of in situ gene expression. Despite rapid development of transcriptome sequencing technologies, which result in the discovery of new candidate genes for many processes, RNA in situ hybridization remains the gold standard for their validation. This review briefly presents modern techniques and their modifications for studying in situ gene expression. Transcriptome studies that have been implemented in cattle, pigs and chickens as model organisms include: in situ RNA hybridization using ZZ-probes, tyramide signal amplification, hybridization chain reaction, digoxigenin-labeled probes, RT-PCR, single cell transcriptome sequencing, in situ transciptome sequencing. This paper is a review of the results of studies on cattle, pigs and chickens. The results of research in this area are relevant for understanding the features of adaptation mechanisms at the transcriptomic level in highly productive animals under industrial conditions in order to search for new markers of valuable agricultural traits. It should be noted that in literature there are very few studies using RNA in situ hybridization, despite the availability and simplicity of the method.

#1945 Comparative analysis of kidney transplantation modeled using precision-cut kidney slices and kidney transplantation in pigs

Abstract Background and Aims Kidney transplants are at risk for so far unavoidable ischemia-reperfusion injury. Several experimental kidney transplantation models are available to study this injury, but all have their own limitations. Here, we describe precision-cut kidney slices (PCKS) as a novel model of kidney ischemia-reperfusion injury in comparison with pig and human kidney transplantation. Method Following bilateral nephrectomy in pigs, we applied warm ischemia (1 h), cold ischemia (20 h) and a reperfusion period (4 h) to one whole kidney undergoing transplantation to a recipient pig and, in parallel, established PCKS undergoing ischemia and modeled reperfusion. Tissues were analyzed using standard histopathology assessment by two blinded pathologists, and bulk RNAseq with gene set enrichment analysis with comparison to human kidney biopsy gene sets obtained from Gene Expression Omnibus. Results Histopathological assessment revealed the presence of some but not all morphological features of tubular injury in PCKS as seen in pig kidney transplantation. RNAseq demonstrated that the majority of changes occurred after reperfusion only, with a partial overlap between PCKS and kidney transplantation, with some differences in transcriptional response attributable to systemic inflammatory responses and immune cell migration. Comparison of PCKS and pig kidney transplantation with RNAseq data from human kidney biopsies by gene set enrichment analysis revealed that both PCKS and pig kidney transplantation reproduced the post-reperfusion pattern of human kidney transplantation. In contrast, only post-cold ischemia PCKS and pig kidney partially resembled the gene set of human acute kidney injury. Conclusion Overall, the present study established that a PCKS protocol can model kidney transplantation and its reperfusion-related damage on a histological and a transcriptomic level. PCKS may thus expand the toolbox for developing novel therapeutic strategies against ischemia-reperfusion injury.

#1493 Upregulated miR-205 as a biomarker of diet-induced obesity-related glomerulopathy

Abstract Background and Aims Obesity is a modifiable risk factor for the development and progression of chronic kidney disease (CKD). Pathways through which obesity might cause renal disease are not completely understood, as not all obese subjects develop CKD. In terms of deepening the knowledge of the preliminary steps of obesity-related renal damage, our group has developed a diet-induced obese murine model with obesity related glomerulopathy (ORG). The objective of this study was to investigate ORG at a cellular, structural and transcriptomic level as a way to discover the triggering pathogenic phenomena of ORG and evaluate potential biomarkers. Method Thirty Wistar rats were randomized into two groups: control rats (n = 15), which were fed with standard diet (SD) and study rats (n = 15), which were fed with high-fat diet (HFD). After 10 weeks, weight, parameters of kidney function, renal histological features, transcriptomic changes, miRNA and mRNA isolation were compared. Results HFD gained significantly more weight (55.8%) than SD(29.2%), p = 0.001. Albuminuria was also significantly higher in HFD (10,384.04) compared with SD (5,845.45), p = 0.001. HFD rats showed typical lesions of early-stages of ORG, with a predominance of mesangial matrix increase (MMI) and podocyte hypertrophy (PH). All histologic lesions correlated with genes differentially expressed (DE) in kidneys of HFD group and, both MMI and PH, also correlated with specific miRNAs DE in the urine of HFD group. The functional analysis showed 4 miRNAs DE in the kidneys of HFD group that negatively regulates PTEN gene, which promotes podocyte endocytosis of lipids in ORG. The electronic microscope confirmed the spaces of lipid vacuoles in the podocytes of HFD rats. Between those 4 miRNAs DE, miR-205 was also found to be upregulated in the urine of HFD group. Conclusion Wistar rats fed with HFD for 10 weeks developed early-stages of ORG, with a specific targetome of miRNAs and gene expression. The upregulation of miR-205 in kidney and its isolation in urine is associated with lipid endocytosis of podocytes, which takes place in the early-stages of ORG and could become a plausible biomarker of early-stages of ORG and open new avenues for future therapeutics research.

#2909 Molecular mechanisms of early vascular damage in children with chronic renal failure

Abstract Background and Aims The risk of developing cardiovascular disease is exceedingly high in patients with chronic kidney disease (CKD), therapeutic options are limited. In order to develop specific therapies, a comprehensive molecular understanding of the underlying pathomechanisms is essential. Method Transcriptome and proteome analyses followed by pathway analyses (GO, GSEA) were performed from microdissected omental arterioles of 6 children with normal renal function and 8 children with CKD5 (median age 9.2 years), respective microproteomics were performed from laser microdissected endothelial layers. Key findings were validated independently by immunohistochemistry. Human endothelial umbilical vein cells (HUVEC) were exposed to plasma from children with normal renal function and children with CKD5. Tight junction (TJ) abundance was quantified in Transwells, ionic conductance by transendothelial resistance measurements and chromatin topology by Single Molecule Localization Microscopy (SMLM) followed by cluster/persistent homology analyses. The latter was compared to ion radiation induced chromatin re-organisation. Results Arteriolar lumen narrowing was already pronounced in children with CKD5 (lumen/vessel ratio; 0.58 vs. 0.76 in healthy individuals, p < 0.001), arteriolar intima and media thickness significantly increased. Biofunctions significantly regulated in CKD compared to controls at both transcriptome and proteome levels were immune response, chromatin organisation, actin cytoskeleton and TJ organization (FDR<0.25, p < 0.05). In independent cohorts of children with CKD5 (n = 20), the subendothelial space was infiltrated by CD68+ macrophages and endothelial histone2A abundance was reduced compared to children with normal renal function (n = 16). Repair foci at the places of DNA double strand breaks (DSB) visualized by yH2A.X were similar in both groups. HUVEC exposed to plasma from CKD5 children showed marked relaxation of H3K9 labeled heterochromatin compared to HUVEC exposed to plasma from children with normal renal function, and persistent homology analysis revealed CKD specific chromatin pattern. The CKD induced chromatin re-organisation differed from ion radiation induced DSB repair. Polarized endothelial cells exposed to plasma from CKD5 children had increased ion transport compared to cells exposed to plasma from healthy donors. Endothelial cell membrane abundance of the sealing TJ protein claudin-5 and of zonula occlundens-1, which links TJ to the actin cytoskeleton, was reduced by uremic plasma, TNFα and carbonyl stress. Co-incubation with histidine dipeptides and AlaGln ameliorated the loss of endothelial barrier function. Conclusion CKD5 leads to complex molecular pathomechanisms and functional alterations particularly including the immune response, chromatin re-organization and endothelial barrier dysfunction already in young children. Dipeptides represent potential therapeutic agents to protect the endothelial barrier function.

#2144 Expression profile induced by TGF-β1 in senescent human peritoneal mesothelial cells is less pronounced than in young cells

Abstract Background and Aims Transforming growth factor β1 (TGF-β1) is a recognized driver of both mesothelial-to-mesenchymal transition (MMT) and cellular senescence observed in vitro in human peritoneal mesothelial cells (HPMCs), and have a significant role in adverse peritoneal remodelling of patients treated with peritoneal dialysis (PD). We have previously shown that senescent HPMCs spontaneously acquire some phenotypic features of MMT which in young HPMCs are induced by TGF-β1. At the same time, senescent HPMCs appeared to be less responsive to TGF-β. It is not clear what determines which pathway is activated by TGF-β1. Here, we aimed to extend previous observations and used electron microscopy, transcriptomics, and proteomics to identify differences between young and senescent cells in their responsiveness to TGF-β. Method Replicative senescence of omentum-derived HPMCs isolated from 4 separate donors was induced by serial passages until cells ceased to grow and expressed senescence-associated β-galactosidase. Serum-starved young and senescent cells were treated in parallel with 1 ng/mL of TGF-β1 for 72 hours. Fixed cells were analyzed with a Jeol transmission electron microscope at 80 kV (Jeol, Tokyo, Japan). Cell lysates were analysed for global gene and protein expression using an integrated transcriptomic (Array-based) and proteomic (LC/MS-based) approach. Results Exposure to TGF-β1 led to phenotypic changes consistent with MMT in young, but not in senescent HPMCs (Fig. 1A). The omics analyses revealed that the response to TGF-β1 was associated with 89 genes being differentially expressed (55 down- and 34 up-upregulated) in young HPMCs and with 13 genes (5 down- and 8 up-regulated) in senescent HPMCs (Fig. 1B). In addition, TGF-β1 changed the expression of 131 proteins in young HPMCs (72 down- and 59 up-regulated) and 232 proteins in senescent HPMCs (127 down- and 105 up-regulated) (Fig. 1C). The responses to TGF-β identified at both the transcriptome and protein levels (Fig. 1D) and present in both young and senescent HPMCs included TGFBI (TGF-β-induced) and ITGB3 (β3 integrin), which were upregulated, and UPK1B (uroplakin 1B) and SEMA3B (semaphorin 3B), which were downregulated. Changes in response to TGF-β seen at the RNA and protein level included also VCAN (versican), ITGA11 (integrin α11), FN1 (fibronectin 1), CCNYL1 (Cyclin Y-like 1) and MYOZ2 (myozenin 2), however, these were observed only in young cells. A decrease in KLK7 (Kallikrein 7) gene expression after exposure to TGF-β was seen both in young and senescent HPMCs, with a decrease in KLK7 protein detected only in senescent cells. Finally, exposure to TGF-β led to an increase in THBS1 (thrombospondin 1) expression in young cells and at the protein level in senescent cells. Conclusion In conclusion, the profound morphological changes seen in senescent HPMCs are not significantly exacerbated by TGFβ. Likewise, changes in global gene and protein profiles in response to TGF-β appear to be less pronounced in senescent cells. Among the genes and proteins affected by TGF-β, those involved in angiogenesis and fibrosis deserve further attention as they may contribute to adverse peritoneal remodelling in PD patients.

Transcriptomic signatures of classical monocytes reveal pro-inflammatory modules and heterogeneity in polyarticular juvenile idiopathic arthritis.

Polyarticular juvenile idiopathic arthritis (pJIA) is a childhood-onset autoimmune disease. Immune cells contribute to persistent inflammation observed in pJIA. Despite the crucial role of monocytes in arthritis, the precise involvement of classical monocytes in the pathogenesis of pJIA remains uncertain. Here, we aimed to uncover the transcriptomic patterns of classical monocytes in pJIA, focusing on their involvement in disease mechanism and heterogeneity. A total of 17 healthy subjects and 18 premenopausal women with pJIA according to ILAR criteria were included. Classical monocytes were isolated, andRNA sequencing was performed. Differential expression analysis was used to compare pJIA patients and healthy control group. Differentially expressed genes (DEGs) were identified, and gene set enrichment analysis (GSEA) was performed.Using unsupervised learning approach, patients were clustered in two groups based on their similarities at transcriptomic level. Subsequently, these clusters underwent a comparative analysis to reveal differences at the transcriptomic level. We identified 440 DEGs in pJIA patients of which 360 were upregulated and 80 downregulated. GSEA highlighted TNF-α and IFN-γ response. Importantly, this analysis not only detected genes targeted by pJIA therapy butalso identified new modulators of immuno-inflammation. PLAUR, IL1B, IL6, CDKN1A, PIM1, and ICAM1 were pointed as drivers of chronic hyperinflammation. Unsupervised learning approach revealed two clusters within pJIA, each exhibiting varying inflammation levels. These findings indicate the pivotal role of immuno-inflammation driven by classical monocytes in pJIA and reveals the existence of two subclusters within pJIA, regardless the positivity of rheumatoid factor and anti-CCP, paving the way to precision medicine.

Metabolic self-feeding in HBV-associated hepatocarcinoma centered on feedback between circulation lipids and the cellular MAPK/mTOR axis

IntroductionHepatitis B Virus (HBV) is widely recognized as a “metabolic virus” that disrupts hepatic metabolic homeostasis, rendering it one of the foremost risk factors for hepatocellular carcinoma (HCC). Except for antiviral therapy, the fundamental principles underlying HBV− and HBV+ HCC have remained unchanged, limiting HCC treatment options.ObjectivesIn this study, we aim to identify the distinctive metabolic profile of HBV-associated HCC, with the promise of identifying novel metabolic targets that confer survival advantages and ultimately impede cancer progression.MethodsWe employed a comprehensive methodology to evaluate metabolic alterations systematically. Initially, we analyzed transcriptomic and proteomic data obtained from a public database, subsequently validating these findings within our test cohort at both the proteomic and transcriptomic levels. Additionally, we conducted a comprehensive analysis of tissue metabolomics profiles, lipidomics, and the activity of the MAPK and AKT signaling pathway to corroborate the abovementioned changes.ResultsOur multi-omics approach revealed distinct metabolic dysfunctions associated with HBV-associated HCC. Specifically, we observed upregulated steroid hormone biosynthesis, primary bile acid metabolism, and sphingolipid metabolism in HBV-associated HCC patients’ serum. Notably, metabolites involved in primary bile acid and sphingolipids can activate the MAPK/mTOR pathway. Tissue metabolomics and lipidomics analyses further validated the serum metabolic alterations, particularly alterations in lipid composition and accumulation of unsaturated fatty acids.ConclusionOur findings emphasize the pivotal role of HBV in HCC metabolism, elucidating the activation of a unique MAPK/mTOR signaling axis by primary bile acids and sphingolipids. Moreover, the hyperactive MAPK/mTOR signaling axis transduction leads to significant reprogramming in lipid metabolism within HCC cells, further triggering the activation of the MAPK/mTOR pathway in turn, thereby establishing a self-feeding circle driven by primary bile acids and sphingolipids.

Toxic mechanisms of the antiviral drug arbidol on microalgae in algal bloom water at transcriptomic level

Increasing antivirals in surface water caused by their excessive consumption pose serious threats to aquatic organisms. Our recent research found that the input of antiviral drug arbidol to algal bloom water can induce acute toxicity to the growth and metabolism of Microcystis aeruginosa, resulting in growth inhibition, as well as decrease in chlorophyll and ATP contents. However, the toxic mechanisms involved remained obscure, which were further investigated through transcriptomic analysis in this study. The results indicated that 885–1248 genes in algae were differentially expressed after exposure to 0.01–10.0 mg/L of arbidol, with the majority being down-regulated. Analysis of commonly down-regulated genes found that the cellular response to oxidative stress and damaged DNA bonding were affected, implying that the stress defense system and DNA repair function of algae might be damaged. The down-regulation of genes in porphyrin metabolism, photosynthesis, carbon fixation, glycolysis, tricarboxylic acid cycle, and oxidative phosphorylation might inhibit chlorophyll synthesis, photosynthesis, and ATP supply, thereby hindering the growth and metabolism of algae. Moreover, the down-regulation of genes related to nucleotide metabolism and DNA replication might influence the reproduction of algae. These findings provided effective strategies to elucidate toxic mechanisms of contaminants on algae in algal bloom water.

Comparative Transcriptomics of Soybean Genotypes with Partial Resistance Toward Phytophthora sojae, Conrad, and M92-220 to Moderately Susceptible Fast Neutron Mutant Soybeans and Sloan.

The breeding of disease-resistant soybeans cultivars to manage Phytophthora root and stem rot caused by the pathogen Phytophthora sojae involves combining quantitative disease resistance (QDR) and Rps gene-mediated resistance. To identify and confirm potential mechanisms of QDR toward P. sojae, we conducted a time course study comparing changes in gene expression among Conrad and M92-220 with high QDR to susceptible genotypes, Sloan, and three mutants derived from fast neutron irradiation of M92-220. Differentially expressed genes from Conrad and M92-220 indicated several shared defense-related pathways at the transcriptomic level but also defense pathways unique to each cultivar, such as stilbenoid, diarylheptanoid, and gingerol biosynthesis and monobactam biosynthesis. Gene Ontology pathway analysis showed that the susceptible fast neutron mutants lacked enrichment of three terpenoid-related pathways and two cell wall-related pathways at either one or both time points, in contrast to M92-220. The susceptible mutants also lacked enrichment of potentially important Kyoto Encyclopedia of Genes and Genomes pathways at either one or both time points, including sesquiterpenoid and triterpenoid biosynthesis; thiamine metabolism; arachidonic acid; stilbenoid, diarylheptanoid, and gingerol biosynthesis; and monobactam biosynthesis. Additionally, 31 genes that were differentially expressed in M92-220 following P. sojae infection were not expressed in the mutants. These 31 genes have annotations related to unknown proteins; valine, leucine, and isoleucine biosynthesis; and protein and lipid metabolic processes. The results of this study confirm previously proposed mechanisms of QDR, provide evidence for potential novel QDR pathways in M92-220, and further our understanding of the complex network associated with QDR mechanisms in soybean toward P. sojae.

SHIP1 modulation and proteome characterization of microglia

Understanding microglial states in the aging brain has become crucial, especially with the discovery of numerous Alzheimer's disease (AD) risk and protective variants in genes such as INPP5D and TREM2, which are essential to microglia function in AD. Here we present a thorough examination of microglia-like cells and primary mouse microglia at the proteome and transcriptome levels to illuminate the roles these genes and the proteins they encode play in various cell states. First, we compared the proteome profiles of wildtype and INPP5D (SHIP1) knockout primary microglia. Our findings revealed significant proteome alterations only in the homozygous SHIP1 knockout, revealing its impact on the microglial proteome. Additionally, we compared the proteome and transcriptome profiles of commonly used in vitro microglia BV2 and HMC3 cells with primary mouse microglia. Our results demonstrated a substantial similarity between the proteome of BV2 and mouse primary cells, while notable differences were observed between BV2 and human HMC3. Lastly, we conducted targeted lipidomic analysis to quantify different phosphatidylinositols (PIs) species, which are direct SHIP1 targets, in the HMC3 and BV2 cells. This in-depth omics analysis of both mouse and human microglia enhances our systematic understanding of these microglia models. SignificanceGiven the growing urgency of comprehending microglial function in the context of neurodegenerative diseases and the substantial therapeutic implications associated with SHIP1 modulation, we firmly believe that our study, through a rigorous and comprehensive proteomics, transcriptomics and targeted lipidomic analysis of microglia, contributes to the systematic understanding of microglial function in the context of neurodegenerative diseases.

A longer time to relapse is associated with a larger increase in differences between paired primary and recurrent IDH wild-type glioblastomas at both the transcriptomic and genomic levels

Glioblastoma (GBM) is the most common malignant brain tumor in adults, which remains incurable and often recurs rapidly after initial therapy. While large efforts have been dedicated to uncover genomic/transcriptomic alternations associated with the recurrence of GBMs, the evolutionary trajectories of matched pairs of primary and recurrent (P-R) GBMs remain largely elusive. It remains challenging to identify genes associated with time to relapse (TTR) and construct a stable and effective prognostic model for predicting TTR of primary GBM patients. By integrating RNA-sequencing and genomic data from multiple datasets of patient-matched longitudinal GBMs of isocitrate dehydrogenase wild-type (IDH-wt), here we examined the associations of TTR with heterogeneities between paired P-R GBMs in gene expression profiles, tumor mutation burden (TMB), and microenvironment. Our results revealed a positive correlation between TTR and transcriptomic/genomic differences between paired P-R GBMs, higher percentages of non-mesenchymal-to-mesenchymal transition and mesenchymal subtype for patients with a short TTR than for those with a long TTR, a high correlation between paired P-R GBMs in gene expression profiles and TMB, and a negative correlation between the fitting level of such a paired P-R GBM correlation and TTR. According to these observations, we identified 55 TTR-associated genes and thereby constructed a seven-gene (ZSCAN10, SIGLEC14, GHRHR, TBX15, TAS2R1, CDKL1, and CD101) prognostic model for predicting TTR of primary IDH-wt GBM patients using univariate/multivariate Cox regression analyses. The risk scores estimated by the model were significantly negatively correlated with TTR in the training set and two independent testing sets. The model also segregated IDH-wt GBM patients into two groups with significantly divergent progression-free survival outcomes and showed promising performance for predicting 1-, 2-, and 3-year progression-free survival rates in all training and testing sets. Our findings provide new insights into the molecular understanding of GBM progression at recurrence and potential targets for therapeutic treatments.

Transcriptome Analysis of Sesame (Sesamum indicum L.) Reveals the LncRNA and mRNA Regulatory Network Responding to Low Nitrogen Stress.

Nitrogen is one of the important factors restricting the development of sesame planting and industry in China. Cultivating sesame varieties tolerant to low nitrogen is an effective way to solve the problem of crop nitrogen deficiency. To date, the mechanism of low nitrogen tolerance in sesame has not been elucidated at the transcriptional level. In this study, two sesame varieties Zhengzhi HL05 (ZZ, nitrogen efficient) and Burmese prolific (MD, nitrogen inefficient) in low nitrogen were used for RNA-sequencing. A total of 3964 DEGs (differentially expressed genes) and 221 DELs (differentially expressed lncRNAs) were identified in two sesame varieties at 3d and 9d after low nitrogen stress. Among them, 1227 genes related to low nitrogen tolerance are mainly located in amino acid metabolism, starch and sucrose metabolism and secondary metabolism, and participate in the process of transporter activity and antioxidant activity. In addition, a total of 209 pairs of lncRNA-mRNA were detected, including 21 pairs of trans and 188 cis. WGCNA (weighted gene co-expression network analysis) analysis divided the obtained genes into 29 modules; phenotypic association analysis identified three low-nitrogen response modules; through lncRNA-mRNA co-expression network, a number of hub genes and cis/trans-regulatory factors were identified in response to low-nitrogen stress including GS1-2 (glutamine synthetase 1-2), PAL (phenylalanine ammonia-lyase), CHS (chalcone synthase, CHS), CAB21 (chlorophyll a-b binding protein 21) and transcription factors MYB54, MYB88 and NAC75 and so on. As a trans regulator, lncRNA MSTRG.13854.1 affects the expression of some genes related to low nitrogen response by regulating the expression of MYB54, thus responding to low nitrogen stress. Our research is the first to provide a more comprehensive understanding of DEGs involved in the low nitrogen stress of sesame at the transcriptome level. These results may reveal insights into the molecular mechanisms of low nitrogen tolerance in sesame and provide diverse genetic resources involved in low nitrogen tolerance research.

Lipid Variability Induces Endothelial Dysfunction by Increasing Inflammation and Oxidative Stress.

This study investigates the impact of fluctuating lipid levels on endothelial dysfunction. Human aortic and umbilical vein endothelial cells were cultured under varying palmitic acid (PA) concentrations: 0, 50, and 100 μM, and in a variability group alternating between 0 and 100 μM PA every 8 hours for 48 hours. In the lipid variability group, cells were exposed to 100 μM PA during the final 8 hours before analysis. We assessed inflammation using real-time polymerase chain reaction, Western blot, and cytokine enzyme-linked immunosorbent assay (ELISA); reactive oxygen species (ROS) levels with dichlorofluorescin diacetate assay; mitochondrial function through oxygen consumption rates via XF24 flux analyzer; and endothelial cell functionality via wound healing and cell adhesion assays. Cell viability was evaluated using the MTT assay. Variable PA levels significantly upregulated inflammatory genes and adhesion molecules (Il6, Mcp1, Icam, Vcam, E-selectin, iNos) at both transcriptomic and protein levels in human endothelial cells. Oscillating lipid levels reduced basal respiration, adenosine triphosphate synthesis, and maximal respiration, indicating mitochondrial dysfunction. This lipid variability also elevated ROS levels, contributing to a chronic inflammatory state. Functionally, these changes impaired cell migration and increased monocyte adhesion, and induced endothelial apoptosis, evidenced by reduced cell viability, increased BAX, and decreased BCL2 expression. Lipid variability induce endothelial dysfunction by elevating inflammation and oxidative stress, providing mechanistic insights into how lipid variability increases cardiovascular risk.

Comprehensive analysis of competing endogenous RNA networks involved in the regulation of glycolysis in clear cell renal cell carcinoma

Abstract Objectives This study aims to elucidate the role of competing endogenous RNAs (ceRNAs), which are pivotal in the post-transcriptional regulation of cancer cells, in the glycolysis of clear cell renal cell carcinoma (ccRCC). Methods RNA-seq data from ccRCC samples and public datasets were subjected to differential expression analysis to identify the upregulated circular RNAs (circRNAs), long noncoding RNAs (lncRNAs), and mRNAs for constructing a ceRNA network. This network focused on the upregulated mRNAs linked to glycolysis and gluconeogenesis, which were verified at the proteome level using the CPTAC database. The ENCORI database was used to predict the lncRNA/circRNA-miRNA and miRNA-mRNA interactions, which formed a network visualized using Cytoscape. This study further examined the association of selected lncRNAs/circRNAs with biological processes and patient survival and explored lncRNA and mRNA expressions at the single-cell level. Results Three circRNAs and eight lncRNAs were identified to be regulators of four glycolysis pathway genes (HK2, LDHA, PFKP, and ALDOC) via 54 miRNAs. Notably, their expressions were aberrant at the transcriptome and proteome levels. These RNA elements were correlated with malignant and immune pathways, and several lncRNAs showed prognostic significance for ccRCC. Furthermore, single-cell RNA sequencing indicated the abnormal enrichment of these lncRNAs and mRNAs in malignant cells. Conclusions This study identified a ceRNA network that plays a critical role in the glycolysis of ccRCC. These findings highlight the importance of ceRNAs in cancer malignancy and their potential impact on patient prognosis.

Toxic mechanisms of nanoplastics exposure at environmental concentrations on juvenile red swamp crayfish (Procambarus clarkii): From multiple perspectives

Nanoplastics pollution has emerged as a global issue due to its widespread potential toxicity. This study delved in to toxic effects of nanoplastics on juvenile P. clarkii and molecular mechanisms from perspectives of growth, biochemical, histopathological analysis and transcriptome level for the first time. The findings of this study indicated that nanoplastics of different concentrations have varying influence mechanisms on juvenile P. clarkii. Nanoplastics have inhibitory effects on growth of juvenile P. clarkii, can induce oxidative stress. The biochemical analysis and transcriptome results indicated that 10 mg/L nanoplastics can activate the antioxidant defense system and non-specific immune system in juvenile P. clarkii, and affect energy metabolism and oxidative phosphorylation. While 20 mg/L and 40 mg/L have a destructive influence on the immune function in juvenile P. clarkii, leading to lipid peroxidation and oxidative damage, and induce apoptosis, can affect ion transport and osmotic pressure regulation. The findings of this study can offer foundational data for delving further into impacts of nanoplastics on crustaceans and toxicity mechanism.